Protective Effect Of Selenium On The Damage Of Liver Cells And Kidney Cells Induced By Sodium Nitrite In Grass Carp(CTENOPHARYNGODON IDELLUS)

There is a gradual decline in the ability of aquatic animals to resist stress,resulting in a gradual increase in aquatic diseases in recent years,due to the expansion of aquaculture,the continuous development of high-density and intensive aquaculture models has been accompanied.It’s a important research that by supplementing some functional nutrients to the fish body,improving the body’s oxidative stress ability and enhancing the body’s immunity in recent years.Selenium has been proven to improve immunity and growth performance in aquatic animals,and is a promising aquatic feed additive.Since the high-density intensive farming mode will greatly increase the nitrite concentration in the aquaculture water,after grass carp supplemented with selenium,there is still little research on the ability to resist high nitrite exposure and its response mechanism.In this study,the grass carp liver and kidney cells were used as the object.From the perspective of selenium and sodium nitrite,the effects of supplementation with appropriate amounts of selenium on the antioxidant and anti-apoptotic capacity of cells under the exposure of sodium nitrite and related intracellular response mechanisms were discussed.The findings are as follows:1. In the grass carp hepatocyte experiment,cells were pre-incubated by Na₂Se O₃ for 12h and then exposed to Na NO₂（5,10,25mg/L）for 24h through the results of CCK8 cell viability test.The results showed that sodium nitrite exposure can inhibit the adhesion of grass carp liver cells,hinder cell growth,the activities of glutathione peroxidase（GPX）,superoxide dismutase（SOD）and catalase（CAT）and the expression of gpx,sod and cat genes were decreased（P<0.05）.The m RNA levels of CHOP（DNA damage-inducing gene 153（GADDl53））and bcl-2 related X protein（bax）genes were up-regulated（P<0.05）,and the apoptotic rate was increased significantly（P<0.05）.After the cells treated by Na₂Se O₃for 24h,the activities of GPX,SOD and CAT was increased（P<0.05）,as well as the expression of gpx,sod,cat,nuclear factor E2 related factor 2（Nrf2）and Kelch-like epichlorohydrin-related protein-1（keap1）genes.When the cells were pre-incubated by Na₂Se O₃for 12h and then exposed to Na NO₂for 24h,the cells could grow normally and the apoptosis rate decreased significantly,and there was no significant difference compared with the control group（P>0.05）,the activities of GPX,SOD,CAT and expression of gpx,sod and cat genes were returned to normal levels（P>0.05）,the expression of B lymphoma-2（bcl-2）gene was significantly up-regulated（P<0.05）.2. In the grass carp kidney cell experiment,cells were pre-incubated by Na₂Se O₃ for 12h and then exposed to Na NO₂（25mg/L）for 24h through the results of CCK8cell viability test.The results showed that the activities of GPX,SOD and CAT was decreased caused by sodium nitrite exposure,in the sodium nitrite exposure group,the m RNA levels of gpx,sod and cat genes were significantly down-regulated（P<0.05）,the expression of chop and bax genes were increased（P<0.05）.The apoptotic rate was increased significantly caused by sodium nitrite exposure.Among them,the intracellular calcium ion concentration increased significantly,and increased with the increase of sodium nitrite concentration,after the BHQ inhibitor incubation,the intracellular calcium ion concentration decreased.In the experimental group exposed to sodium nitrite after incubation with sodium selenite,both enzyme activity-related genes and apoptosis-related genes could be maintained at normal levels,and the intracellular calcium ion concentration was not significantly different from that of the control group（P>0.05）.3. In summary,the pre-incubation of grass carp liver and kidney cells with sodium selenite can effectively alleviate the oxidative damage caused by sodium nitrite exposure,reduce the apoptosis rate,and maintain the normal growth of cells;When the cells were pre-incubated by Na₂Se O₃,the expression of antioxidant genes and key genes in the Nrf2-keap1 pathway were increased significantly,suggesting that the protective effect of selenium incubation may be achieved through the Nrf2-keap1pathway.In kidney cells,the pre-incubation of selenium can alleviate the intracellular endoplasmic reticulum calcium ion disorder,indicating that in reducing the rate of apoptosis,it may be related to the alleviation of intracellular calcium ion disorder.